LOCUS       sequence-160586-        4769 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  fission yeast epitope tagging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4769)
  AUTHORS   Amelina H, Moiseeva V, Collopy LC, Pearson SR, Armstrong CA, Tomita 
            K
  TITLE     Sequential and counter-selectable cassettes for fission yeast.
  JOURNAL   BMC Biotechnol. 2016 Nov 8;16(1):76.
  PUBMED    27825338
REFERENCE   2  (bases 1 to 4769)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "BMC 
            Biotechnol."; date: "2016-11-8"; volume: "16(1)"; pages: "76"
COMMENT     Sequence Label: pNX3b-PK12
FEATURES             Location/Qualifiers
     source          1..4769
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     43..61
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(99..121)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     221..240
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        321..339
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     321..338
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             406..447
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             451..492
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             493..534
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             541..582
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             586..627
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             628..669
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             676..717
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             721..762
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             763..804
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             811..852
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             856..897
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             898..939
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     terminator      969..1156
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1024..1048)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1215..2790
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        1215..1558
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1559..2587
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLMHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     terminator      2593..2790
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2849..2866)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3020..3608)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3100..3119)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3779..4570,4571..4639))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4402..4421
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4640..4744)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg
       61 ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg
      121 gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg
      181 tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta
      241 ctgagagtgc accatatgga catattgtcg ttagaacgcg gctacaatta atacataacc
      301 ttatgtatca tacacatacg atttaggtga cactatagaa cgcggccgcc agctgaagct
      361 tcgtacgctg caggtcgacg gatccccggg ttaattaacg ctagcggtaa gcctattcct
      421 aaccctcttt tgggtctcga ttctacaggt ggaaaaccaa tccccaaccc actcctcggc
      481 cttgactcaa ctggcaaacc cataccaaat cctttacttg gattagatag tacctctagc
      541 ggtaagccta ttcctaaccc tcttttgggt ctcgattcta caggtggaaa accaatcccc
      601 aacccactcc tcggccttga ctcaactggc aaacccatac caaatccttt acttggatta
      661 gatagtacct ctagcggtaa gcctattcct aaccctcttt tgggtctcga ttctacaggt
      721 ggaaaaccaa tccccaaccc actcctcggc cttgactcaa ctggcaaacc cataccaaat
      781 cctttacttg gattagatag tacctctagc ggtaagccta ttcctaaccc tcttttgggt
      841 ctcgattcta caggtggaaa accaatcccc aacccactcc tcggccttga ctcaactggc
      901 aaacccatac caaatccttt acttggatta gatagtacct ctagatgagg cgcgccactt
      961 ctaaataagc gaatttctta tgatttatga tttttattat taaataagtt ataaaaaaaa
     1021 taagtgtata caaattttaa agtgactctt aggttttaaa acgaaaattc ttattcttga
     1081 gtaactcttt cctgtaggtc aggttgcttt ctcaggtata gtatgaggtc gctcttattg
     1141 accacacctc taccggcaga tccgctagga gatctgttta gcttgcctcg tccccgccgg
     1201 gtcacccggt cagcgacatg gaggcccaga ataccctcct tgacagtctt gacgtgcgca
     1261 gctcaggggc atgatgtgac tgtcgcccgt acatttagcc catacatccc catgtataat
     1321 catttgcatc catacatttt gatggccgca cggcgcgaag caaaaattac ggctcctcgc
     1381 tgcggacctg cgagcaggga aacgctcccc tcacagacgc gttgaattgt ccccacgccg
     1441 cgcccctgta gagaaatata aaaggttagg atttgccact gaggttcttc tttcatatac
     1501 ttccttttaa aatcttgcta ggatacagtt ctcacatcac atccgaacat aaacaaccat
     1561 gggtaaaaag cctgaactca ccgcgacgtc tgtcgagaag tttctgatcg aaaagttcga
     1621 cagcgtctcc gacctgatgc agctctcgga gggcgaagaa tctcgtgctt tcagcttcga
     1681 tgtaggaggg cgtggatatg tcctgcgggt aaatagctgc gccgatggtt tctacaaaga
     1741 tcgttatgtt tatcggcact ttgcatcggc cgcgctcccg attccggaag tgcttgacat
     1801 tggggaattc agcgagagcc tgacctattg catctcccgc cgtgcacagg gtgtcacgtt
     1861 gcaagacctg cctgaaaccg aactgcccgc tgttctgcag ccggtcgcgg aggccatgga
     1921 tgcgatcgct gcggccgatc ttagccagac gagcgggttc ggcccattcg gaccgcaagg
     1981 aatcggtcaa tacactacat ggcgtgattt catatgcgcg attgctgatc cccatgtgta
     2041 tcactggcaa actgtgatgg acgacaccgt cagtgcgtcc gtcgcgcagg ctctcgatga
     2101 gctgatgctt tgggccgagg actgccccga agtccggcac ctcatgcacg cggatttcgg
     2161 ctccaacaat gtcctgacgg acaatggccg cataacagcg gtcattgact ggagcgaggc
     2221 gatgttcggg gattcccaat acgaggtcgc caacatcttc ttctggaggc cgtggttggc
     2281 ttgtatggag cagcagacgc gctacttcga gcggaggcat ccggagcttg caggatcgcc
     2341 gcggctccgg gcgtatatgc tccgcattgg tcttgaccaa ctctatcaga gcttggttga
     2401 cggcaatttc gatgatgcag cttgggcgca gggtcgatgc gacgcaatcg tccgatccgg
     2461 agccgggact gtcgggcgta cacaaatcgc ccgcagaagc gcggccgtct ggaccgatgg
     2521 ctgtgtagaa gtactcgccg atagtggaaa ccgacgcccc agcactcgtc cgagggcaaa
     2581 ggaataatca gtactgacaa taaaaagatt cttgttttca agaacttgtc atttgtatag
     2641 tttttttaca ttgtagttgt tctattttaa tcaaatgtta gcgtgattta tatttttttt
     2701 cgcctcgaca tcatctgccc agatgcgaag ttaagtgcgc agaaagtaat atcatgcgtc
     2761 aatcgtatgt gaatgctggt cgctatactg ctgtcgattc gatactaacg ccgccatcca
     2821 gtttaaacga gctcgaattc taggcttcct cgctcactga ctcgctgcgc tcggtcgttc
     2881 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     2941 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     3001 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     3061 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     3121 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     3181 cctttctccc ttcgggaagc gtggcgcttt ctcaatgctc acgctgtagg tatctcagtt
     3241 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     3301 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     3361 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     3421 agttcttgaa gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg
     3481 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     3541 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     3601 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     3661 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     3721 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     3781 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     3841 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     3901 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     3961 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     4021 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     4081 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     4141 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     4201 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     4261 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     4321 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     4381 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     4441 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     4501 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     4561 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     4621 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     4681 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     4741 cgcgcacatt tccccgaaaa gtgccacct
//
